The pathological hallmarks of Alzheimer's disease (AD) are the presence of senile plaques, largely composed of extracellular deposits of amyloid beta (A2) peptide and intracellular neurofibrillary tangles. Increasing evidence suggests that the low density lipoprotein receptor-related protein (LRP) facilitates amyloidogenic processing of APP. Within LRP, we defined the last 37 C-terminal residues of LRP (LRP-C37) lacking the NPxY or YxxL domains sufficient to robustly promote A2 production. The role of LRP-C37 domain in LRP function is largely unknown. Since LRP-C37 alone was a potent inducer of A2 production, we used this domain as bait in a yeast 2-hybrid screen, resulting in the identification of Ran-binding protein M (RanbpM), implicated in intracellular membrane trafficking and signaling. RanbpM interacted with both LRP and APP in transfected cells and mouse brain homogenates. Over-expression of RanbpM also enhanced the amount of LRP/APP complex formation in mammalian cells, suggesting a role for RanbpM as an adaptor protein that facilitates the tripartite interaction among APP, RanbpM and LRP. When 90 kDa full-length RanbpM (RanbpM-FL) was over-expressed in stable cell lines, a proteolytically cleaved N-terminal 55 kDa fragment (N-55) was also generated, which interacted strongly with both APP and LRP even more than the full-length RanbpM. This proteolytically cleaved species of RanbpM was elevated by 6-fold in brains of AD patients. Indeed, over-expression of RanbpM-FL or the N55 fragment markedly increased A2 secretion. We hypothesize that RanbpM alters APP metabolism in vivo, in particular via the N55 fragment of RanbpM. The specific aims of this application are to first generate transgenic mice with neuronal expression of RanbpM-FL and RanbpM-N55 and then to cross with mice over-expressing APP to obtain RanbpM/APP double transgenic mice. Whether over-expression of RanbpM variants alter amyloidogenic processing of APP will be determined by quantifying the levels of A2 and amyloid plaques by biochemical and immunohistochemical examinations in the cortex and hippocampus of Tg RanbpM/APP double transgenic mice. The level of CHAPS-soluble and insoluble A2 will be quantified by ELISA at 2 and 10 months of age. The characterization of novel APP and LRP-interacting proteins that promote amyloidogenic processing of APP will reveal novel sites of pathogenesis and targets for anti-amyloid intervention. We believe that RanbpM represents a potential therapeutic target. In addition, the RanbpM transgenic mice generated through this R03 granting mechanism is expected to hold substantial extrinsic merit, as they will be undoubtedly be useful to many researchers who study other aspects of RanbpM. This application may also lay the foundation for a future R01 investigation on the role of RanbpM in neurodegeneration. PUBLIC HEALTH RELEVANCE: Alzheimer's disease (AD), a disease characterized by toxic amyloid (A2) accumulation in brain, affects about 5 million people in America, but despite extensive research, available treatments provide only limited symptomatic relief. Recently we found a protein called RanbpM that enhances the production of amyloid/A2 in cells cultured in the laboratory. In this application, we will express RanbpM in brains of mice to confirm the findings in animal models in the hopes of that new insights may be uncovered for therapeutically targeting amyloid production in AD, in particular with RanbpM as a therapeutic target in mind.